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AB0026 Pathophysiological hypoxia affects both redox state and il-2 signaling of human cd4+ t cells and concomitantly impairs survival and proliferation
  1. T. Gaber1,2,3,
  2. C. L. Tran1,2,
  3. S. Schellmann1,2,
  4. M. Hahne1,2,4,
  5. C. Strehl1,2,
  6. M. Jakstadt1,2,3,
  7. G.-R. Burmester1,
  8. F. Buttgereit1
  1. 1Department of Rheumatology and Clinical Immunology, Charité University Hospital
  2. 2German Rheumatism Research Center (DRFZ)
  3. 3Berlin-Brandenburg Center of Regenerative Therapies (BCRT)
  4. 4Berlin-Brandenburg School of Regenerative Therapies (BSRT), Berlin, Germany

Abstract

Background Rheumatoid arthritis (RA) is a chronic inflammatory disease of diarthrodial joints, characterized by the infiltration of immune cells, alterations of cellular redox state and pathophysiological local tissue hypoxia (<1% O2).

Objectives We investigated the impact of different hypoxia levels on survival, proliferation, cytokine secretion, intracellular energy and redox state of quiescent and mitogen stimulated human CD4+ T cells.

Methods Isolated human CD4+ T cells were exposed to normoxia (18% O2), physiological hypoxia (5% O2) and pathophysiological hypoxia (<1% O2), respectively and cultured with or without PHA stimulation at defined periods of time (6h, 24, 48, and 72h). Cells were then analyzed by measuring IL-2R signaling (CD25 expression, phosphoSTAT5a), proliferation and the production of intracellular ROS (iROS) using flow cytometry. In addition, caspase-3/7 activity and ATP levels were determined by luminometric assays. Cellular oxidative damage and cytokine release were analyzed by ELISA based quantification of protein carbonylation (ROS damage) and IL2 production.

Results We found pathophysiological hypoxia (<1% O2) to decrease significantly CD4+ T cell survival after mitogenic stimulation. This effect was neither due to induced caspase-3/7-mediated apoptosis nor to increased ATP consumption or increased ATP depletion. However, the ability of stimulated T cells to proliferate was both delayed and suppressed under these hypoxic conditions, despite increased expression of CD25 and unchanged secreted IL-2 amounts. Hypoxia was also found to modify iROS levels in stimulated T cells over time, but without producing measurable effects on oxidative protein damage as compared to normoxia. In contrast, both normoxia (18% O2) and physiological hypoxia (5% O2) did not decrease CD4+ T cell survival and proliferation after mitogenic stimulation.

Conclusions We conclude that pathophysiological hypoxia (<1% O2) but not physiological hypoxia (5% O2) affects T cell proliferation and viability via disturbed IL-2R signaling downstream of STAT5a phosphorylation. However, the decreased T cell proliferation was not linked to an impaired cellular energy homeostasis. We suggest iROS links early events in T cell stimulation to the inhibition of the lymphoproliferative response under hypoxic conditions. The level of iROS may therefore act as a mediator of immune functions leading to down-regulation of long-term T cell activity in inflamed tissues.

Disclosure of Interest None Declared

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